【摘要】：第一部分成年期追趕生長大鼠模型的建立及一般生物學(xué)特征觀察 目的：建立成年期追趕生長大鼠模型并觀察其一般生物學(xué)特征。 方法：采用體重因素隨機(jī)區(qū)組設(shè)計,按體重區(qū)組將6周齡雄性SD大鼠24只隨機(jī)分為4組(共2個時間點(diǎn)),4周時間點(diǎn)2組：熱卡限制4周組(R4),正常飲食4周組(NC4)；12周時間點(diǎn)2組：正常飲食追趕生長8周組(RN8)和持續(xù)正常飲食12周組(NC12)。通過先熱卡限制后恢復(fù)飲食的方法建立成年期追趕生長大鼠模型。實(shí)驗(yàn)過程中每天記錄實(shí)驗(yàn)大鼠進(jìn)食量及體重變化,實(shí)驗(yàn)結(jié)束時觀察實(shí)驗(yàn)大鼠熱卡攝入量、體重和雙能X線吸收測量法DEXA (Dual Energy X-ray Absorptiometry)檢測不同部位的脂肪含量。高胰島素-正糖鉗夾實(shí)驗(yàn)評估追趕生長大鼠系統(tǒng)胰島素敏感性及血糖、血清胰島素水平檢測。 結(jié)果： 1、實(shí)驗(yàn)動物實(shí)際能量攝入在限食期間約為NC組的60%,恢復(fù)飲食后基本達(dá)到NC組水平。 2、RN組在熱卡限制期生長受抑,體重明顯下降�；謴�(fù)飲食后,體重增加迅速,但實(shí)驗(yàn)結(jié)束時仍未達(dá)NC組水平。 3、DEXA檢測結(jié)果示限食時軀干組織脂肪含量及軀干脂肪占全身脂肪比例較正常對照組均下降,開放飲食后,RN8組全身、軀干脂肪含量及軀干脂肪比例均顯著高于NC組。 4、高胰島素-正葡萄糖鉗夾實(shí)驗(yàn)顯示RN組存在嚴(yán)重的系統(tǒng)胰島素抵抗。R4組血糖和胰島素水平較NC4組無明顯變化,RN8組血糖水平較NC12組變化不明顯,但胰島素水平顯著升高。 結(jié)論：通過熱卡限制后恢復(fù)飲食方法建立的成年期追趕生長大鼠模型表現(xiàn)為熱卡限制期體重生長抑制,恢復(fù)飲食后體重增長迅速和明顯的體脂含量及內(nèi)臟脂肪比例升高,且存在顯著地系統(tǒng)胰島素抵抗。與人群中發(fā)生的追趕生長時所觀察到的變化類似,可作為一種較為理想的研究追趕生長的模型。 目的：探討成年期追趕生長脂肪組織脂質(zhì)生成與儲存能力的改變和胰島素抵抗之間的可能機(jī)制。 方法：采用體重因素隨機(jī)區(qū)組設(shè)計,按體重區(qū)組將6周齡雄性SD大鼠隨機(jī)分為8組其中4組(即追趕生長組群RN)為熱卡限制組(R4)和追趕生長2、4、8周組(熱卡限制后開放正常飲食RN2、RN4、RN8)；另外4組為與之年齡匹配的正常飲食對照組(即NC組群,分別為NC4、NC6、NC8、NC12)。檢測附睪和皮下脂肪組織PPAR-γ、 FSP27的表達(dá)水平,檢測兩種脂肪組織在熱卡限制期末及追趕生長8周結(jié)束時(R4、RN8)兩個時間點(diǎn)的高胰島素-正糖鉗夾實(shí)驗(yàn)過程中2-脫氧-D葡萄糖攝取、血清甘油三酯、游離脂肪酸、脂聯(lián)素及腫瘤壞死因子-α水平。 結(jié)果： 1、與NC4組相比,R4組附睪脂肪PPAR-γ的表達(dá)顯著增加,而FSP27的表達(dá)明顯下降；R4組皮下脂肪組織PPAR-γ的表達(dá)亦顯著增加,但FSP27無明顯變化。 2、與NC4組相比,R4組附睪脂肪和皮下脂肪組織2-脫氧-D葡萄糖攝取率輕度增加,但均未達(dá)到統(tǒng)計學(xué)意義。 3、R4組血清游離脂肪酸、脂聯(lián)素水平較NC4組明顯升高,甘油三酯、腫瘤壞死因子-α變化不顯著。 4、附睪脂肪組織的PPAR-γ的表達(dá)水平在RN2、RN4、RN8組均明顯增加,FSP27的mRNA表達(dá)水平在RN2組低于NC6組,在RN4組與NC8組無統(tǒng)計學(xué)差異,在RN8組明顯高于NC12組。 5、皮下脂肪組織的PPAR-γ和FSP27表達(dá)水平在RN2、RN4、RN8組均無明顯差異。 6、RN8組附睪脂肪組織2-脫氧-D葡萄糖攝取較NC12組顯著增加,而皮下脂肪組織變化不明顯。 7、RN8組血清游離脂肪酸、甘油三酯、腫瘤壞死因子-α水平均顯著升高,而脂聯(lián)素水平則明顯降低。 結(jié)論：限食階段內(nèi)臟脂肪組織脂質(zhì)生成和儲存能力改變的不一致導(dǎo)致恢復(fù)飲食后脂質(zhì)儲存能力相對不足,加之生脂原料增加,使得過多的脂質(zhì)不能有效的儲存,脂質(zhì)外溢,炎癥因子釋放增加,最終導(dǎo)致系統(tǒng)胰島素抵抗的形成。皮下脂肪組織在該過程中變化不顯著。因此,內(nèi)臟脂肪的脂質(zhì)生成和儲存能力的失衡可能是成年期追趕生長胰島素抵抗形成的重要原因。 目的：評估白藜蘆醇在成年期追趕生長大鼠模型中對脂肪組織脂質(zhì)生成和儲存能力的影響。 方法：分組方法同第一部分,在4周和12周時間點(diǎn)各增加1組分別為R4+白藜蘆醇、RN8+白藜蘆醇(R4E、RN8E)組,即選取熱卡限制結(jié)束(R4)及恢復(fù)飲食追趕生長結(jié)束(RN8)兩個時間點(diǎn)作為觀察白藜蘆醇干預(yù)效果的研究。每天進(jìn)行一定劑量白藜蘆醇灌胃治療,其余4組(NC4、R4、NC12和RN8)均進(jìn)行相同劑量生理鹽水灌胃治療。記錄白藜蘆醇干預(yù)組的體重,DEXA檢測干預(yù)組體脂含量,高胰島素-正糖鉗夾實(shí)驗(yàn)評估白藜蘆醇對追趕生長大鼠(R4E和RN8E組)系統(tǒng)胰島素敏感性的影響；檢測4周組和8周組干預(yù)前后附睪和皮下脂肪組織SIRT-1的表達(dá)水平及活性；檢測干預(yù)后附睪和皮下脂肪組織PPAR-γ和FSP27的表達(dá)水平、高胰島素-正糖鉗夾實(shí)驗(yàn)過程中2-脫氧-D葡萄糖攝取率、脂肪細(xì)胞的形態(tài)學(xué)改變；檢測12周組附睪脂肪組織AKT及磷酸化AKT的表達(dá)；檢測干預(yù)后血清甘油三酯、游離脂肪酸、脂聯(lián)素及腫瘤壞死因子-α水平。 結(jié)果： 1、R4E和RN8E組全身組織脂肪含量、軀干組織脂肪含量、軀干脂肪占全身脂肪比例,均較干預(yù)前明顯下降,達(dá)到甚至低于NC組。體重較干預(yù)前僅輕度下降,與RN組相比無統(tǒng)計學(xué)意義。 2、白藜蘆醇對R4E組系統(tǒng)胰島素敏感性、空腹血糖及空腹胰島素水平無明顯影響；RN8E組胰島素敏感性和空腹胰島素顯著改善,基本恢復(fù)至正常水平,但對血糖水平影響不明顯。 3、4周組中SIRT-1的表達(dá)和活性與NC4組相比,兩種脂肪組織在R4組均明顯升高；在12周組中與NC12組相比,兩種脂肪組織在RN8組均明顯降低；白藜蘆醇干預(yù)后,兩種組織在R4E、RN8E組SIRT-1的表達(dá)和活性超過或達(dá)到正常對照組。 4、經(jīng)白藜蘆醇干預(yù)后,附睪脂肪組織R4E、RN8E組FSP27的mRNA表達(dá)水平較干預(yù)前顯著增加,超過NC組；PPAR-γ的表達(dá)明顯下降,接近或低于NC組；2-脫氧-D葡萄糖攝取顯著增加,高于NC組；RN8脂肪細(xì)胞組形狀不規(guī)則,直徑變大,胞膜不完整；RN8E組脂肪細(xì)胞形態(tài)學(xué)趨于正常。 5、與NC12組相比,RN8組大鼠附睪脂肪Ser473PKB磷酸化程度無顯著變化,Thr308PKB磷酸化程度明顯升高；白藜蘆醇干預(yù)后,Ser473和Thr308PKB磷酸化程度均明顯高于NC12及RN8組。 6、經(jīng)白藜蘆醇干預(yù)后,皮下脂肪組織R4E、RN8E組PPAR-γ的表達(dá)水平較干預(yù)前顯著降低,接近或低于NC組；2-脫氧-D葡萄糖攝取顯著增加,高于NC組；脂肪細(xì)胞在RN8、RN8E組形態(tài)學(xué)變化不明顯。 7、R4E組和RN8E組血清游離脂肪酸、甘油三酯、腫瘤壞死因子-α均較干預(yù)前有所下降,在RN8E組下降趨勢更為明顯；脂聯(lián)素水平則較干預(yù)前顯著升高,恢復(fù)至NC組水平。 結(jié)論：白藜蘆醇可抑制追趕生長過程中脂質(zhì)的生成能力,提升脂質(zhì)的儲存能力,在一定程度上使二者改變接近平衡,并且保護(hù)脂肪細(xì)胞的完整性,減少脂質(zhì)外溢及炎癥因子釋放,從而從本質(zhì)上改善追趕生長所致的系統(tǒng)胰島素抵抗。
[Abstract]:The establishment and general biological characteristics of the model of the first part after-one-year follow-up growth in rats Objective: To establish a rat model of catching and growing in adulthood and to observe its general biology. Methods: The 24-week-old male SD rats were randomly divided into 4 groups (2 time points) and 4 weeks at the time of 4 weeks:4 weeks (4 weeks),4 weeks (NC4) and 12 weeks in the body weight group. Point 2: normal diet followed up for 8 weeks (RN8) and continued normal diet for 12 weeks (NC 12) The method of recovering the diet after the restriction of the thermal card is established, and the growth of the one-year after-catch growth is established. Rat model was used to study the changes of feeding amount and body weight of the experimental rats per day during the course of the experiment. The experiment was conducted to observe the caloric intake, body weight and dual energy X-ray absorptiometry (DEXA (Dual Energy X-ray Absorbance) of the experimental rats. Determination of insulin sensitivity and blood glucose and serum insulin water in rats after catching and growing in rats with high insulin-positive sugar flat detection Results:1. The actual energy intake of the experimental animals was about 60% of the NC group during the food-limiting period. The level of the NC group is reached.2, the RN group is grown during the thermal card limit period. The body weight was significantly reduced. After the recovery of the diet, the weight gain was rapid, but the experiment was over The level of the NC group was still not reached.3. The results of DEXA test showed that the body fat content and the body fat in the body were lower than that in the normal control group. After the open diet, the body fat content and the trunk fat of the whole body, trunk and body of the RN8 group were decreased. The ratio of the two groups was significantly higher than that of the NC group.4. The experiment of high insulin-positive glucose clamp showed that the RN There was a serious systemic insulin resistance in the group. There was no significant change in the blood glucose and insulin levels in the group R4, and the level of blood glucose in the RN8 group was less than that of the N12 group. The results showed that the growth of body weight and body fat content and the proportion of visceral fat increased rapidly and the body fat content of the rats after the recovery of the diet was increased rapidly and the body fat content and the visceral fat ratio were increased after the diet was restored by the hot-card restriction. high, and there is a significant system insulin resistance, similar to that observed in the follow-up growth that occur in the population, An ideal model for the study of the catch-up and growth of adipose tissue was studied. Objective: To study the formation and storage of lipid in the growth adipose tissue during the period of adult-year follow-up. Methods:6-week-old male SD rats were randomly divided into 8 groups by weight-factor random-zone design,4 groups (i.e., catch-up group RN) were heat-card-limiting group (R4) and after-catch growth of 2,4, and 8-week group (thermal card limitation). The normal diet (RN2, RN4, RN8) and the normal diet control group (i.e., NC group, sub-group) matched with the age of the normal diet were group 4. The expression levels of PPAR-1 and FSP27 in the epididymis and the subcutaneous fat tissue were detected, and the high insulin-positive ratio of the two adipose tissues at two time points at the end of the caloric restriction period and at the end of the after-catch growth of 8 weeks (R4, RN8) was detected. 2-deoxy-D glucose uptake, serum triglyceride, and swim in the course of the sugar-jaw experiment off fat Results:1. The expression of PPAR-1 in the epididymis of R4 group increased significantly compared with that of the NC4 group, while the expression of FSP27 decreased significantly. The expression of R-1 was also significantly increased, but there was no significant change in FSP27. In comparison with the NC4 group, the epididymal fat and the subcutaneous fat tissue of the group R4 were 2- The results showed that the levels of free fatty acid and adiponectin in group 3 and R4 were higher than those in group 3 and 4. The expression of PPAR-1 in the adipose tissue of the epididymis increased significantly in the RN2, RN4 and RN8 groups, and the expression level of FSP27 was lower than that of the NC6 group in the RN2 group. There was no statistical difference between the N4 group and the NC8 group, and the PPAR of the subcutaneous fat tissue was significantly higher in the RN8 group than in the NC12 group. The expression levels of-1 and FSP27 were not significantly different in RN2, RN4 and RN8 groups. -Deoxy-D glucose uptake increased significantly in the NC12 group, while the changes in the subcutaneous fat tissue were not significant. Conclusion: The changes of lipid production and storage capacity of visceral fat tissue in the limited-feeding stage are not consistent with the change of lipid production and storage capacity of the visceral fat tissue in the limiting stage, and the lipid storage capacity after the recovery of the diet is relatively low. In addition, the raw material of the raw fat is increased, so that the excessive fat It is not effective in storage, lipid spillovers, and inflammation. The increase of the sub-release results in the formation of the system's insulin resistance. The subcutaneous fat tissue changes in this process. Therefore, the visceral organs Imbalance of the lipid generation and storage capacity of fat may be an important cause of the formation of the growth of insulin resistance in adulthood. Objective: To evaluate the effect of the white-calcitol on the lipid formation and storage capacity of adipose tissue in the growth rat model after the adulthood. Methods: The grouping method is the same as the first part, and the effect of the method is 4 weeks and 4 weeks. At the 12-week time point, group 1 was the group R4 + white, and the group of R4E and RN8E respectively, that is, the heat-card-limiting junction was selected. A study of the effect of the two time points of the beam (R4) and the recovery diet after the end of the growth (RN8) was used as a study to observe the effect of the intervention of the aloe. The rats were treated with the same dose of saline in the other four groups (NC4, R4, NC12, and RN8). To evaluate the effect of the white-calcitol on the insulin sensitivity of the rats (R4E and RN8E) in the follow-up growth of rats (R4E and RN8E). The expression level and activity of SIRT-1 in the epididymis and the subcutaneous fat tissue before and after the intervention of the 4-week and 8-week groups were examined. The expression level of R-1 and FSP27, the rate of 2-deoxy-D glucose uptake and the morphological changes of fat cells during the experiment of high insulin-positive sugar clamp, and the detection of AKT and P in the epididymis adipose tissue in the 12-week group. Acidification A Expression of KT, serum triglyceride, free fatty acid, adiponectin and tumor necrosis factor-1 level after intervention. Results:1, R4E and RN8E were all body tissues. Fat content, body fat content, body fat proportion of body fat There was no statistical significance in the insulin sensitivity, fasting blood glucose and fasting insulin levels in the R4E group. The expression and activity of SIRT-1 in group 3 and 4 were significantly higher in group 4 than in the NC4 group. In the 12-week group, the two adipose tissues were in the RN8 group compared with the N12 group. The expression and activity of the two tissues in the R4E and RN8E group SIRT-1 exceeded or reached the normal control group. And the expression of PPAR-1 is significantly lower, close to or lower than that of NC group, and 2-deoxy-D glucose uptake is obvious. The increase was higher than that of the NC group, the shape of the group of RN8 adipocytes was irregular, the diameter of the cell was large, the cell membrane was not complete, the morphology of the fat cells in the RN8E group tended to be normal.5. Compared with the NC12 group, there was no significant change in the phosphorylation of the fat Ser473PKB in the epididymis of the RN8 group, Thr308PKB The level of phosphorylation of Ser473 and Thr308PKB was significantly higher than that of NC12 and RN8 groups. Low, close to or lower than NC group;2-deoxy-D glucose uptake increased significantly, which was higher than that of NC group; the morphological changes of fat cells in RN8 and RN8E group were not obvious. And the level of adiponectin decreased significantly before intervention and returned to the level of NC group. The ability to produce, to improve the storage capacity of the lipid, to a certain extent
【學(xué)位授予單位】：華中科技大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2012
【分類號】：R285.5

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